Flap endonuclease 1 (FEN1) has been shown to remove 5 overhanging flap intermediates during base excision repair and to process the 5 ends of Okazaki fragments during lagging-strand DNA replication in vitro. To assess the in vivo role of the mammalian enzyme in repair and replication, we used a gene-targeting approach to generate mice lacking a functional Fen1 gene. Heterozygote animals appear normal, whereas complete depletion of FEN1 causes early embryonic lethality. Fen1 ؊/؊ blastocysts fail to form inner cell mass during cellular outgrowth, and a complete inactivation of DNA synthesis in giant cells of blastocyst outgrowth was observed. Exposure of Fen1 ؊/؊ blastocysts to gamma radiation caused extensive apoptosis, implying an essential role for FEN1 in the repair of radiation-induced DNA damage in vivo. Our data thus provide in vivo evidence for an essential function of FEN1 in DNA repair, as well as in DNA replication.Flap endonuclease 1 (FEN1) is a member of the RAD2 superfamily of nucleases, which play a critical role in DNA replication and repair in both prokaryotes and eukaryotes (16,17,24,26,55). The main function of FEN1 in replication is proposed to be the removal of displaced RNA-DNA primers synthesized by DNA polymerase ␣-primase during discontinuous lagging-strand replication. A similar single-stranded DNA flap structure is produced by excessive gap filling by a DNA repair polymerase during long-patch base excision repair (BER). Both biochemical and genetic studies support a role for FEN1 during these cellular processes. Similar 5Ј-flap intermediates are also formed in nonhomologous DNA end joining of double-strand DNA breaks and DNA recombination (25,57).The biochemistry of FEN1 has been studied extensively by several groups (reviewed in reference 32), and the crystal structures of two FEN1 orthologues from archaea have been solved (19,20). The enzyme employs a unique cleavage mechanism for substrates containing single-stranded 5Ј tails or 5Ј-flap structures. It recognizes the 5Ј end, tracking the length of the tail, and cleaves at the junction between double-stranded and single-stranded DNAs (40). Although FEN1 acts less efficiently as an exonuclease than as a flap endonuclease, it is likely that the enzyme employs similar mechanisms for both reactions (16,34).FEN1 has been shown to have an important role in the processing of intermediates formed during BER of modified structures in DNA. Recent evidence has indicated that in mammalian cells, BER is mediated through at least two subpathways with different repair patch sizes and different enzymes involved. These pathways have been designated singlenucleotide BER and long-patch BER (12,27,28). The choice of subpathways in BER depends on whether the 5Ј deoxyribose phosphate intermediate can be efficiently removed by the polymerase ␤ lyase activity to yield a 5Ј-phosphorylated DNA strand capable of serving as a substrate for DNA ligase (44). When such processing is inefficient, long-patch BER can occur, and FEN1 will remove the overhang formed by re...

OGG1 (8-oxoguanine DNA glycosylase-1) is one of the main DNA glycosylases present in mammalian cells. The enzyme removes 7,8-dihydro-8-oxoguanine (8-oxoG) lesions, believed to be the most important oxidized lesions due to their relatively high incidence and their miscoding properties. This study shows that in prenatal mice brains the repair capacity for 8-oxoG is 5-10-fold higher than in adult mice brains. Western blot analysis and repair activity in extracts from Ogg1 À/À mice revealed that OGG1 was responsible for the efficient 8-oxoG removal from prenatal mice. To investigate how OGG1 protects against oxidative stress-induced mutagenesis, pregnant Big Blue/wild-type and Big Blue/Ogg1 À/À mice were exposed to nontoxic doses of gamma radiation. A 2.5-fold increase in the mutation frequency in Ogg1 À/À mouse brains was obtained by exposure to 3.5 Gy at day 19 postfertilization. This was largely due to GC to TA transversions, believed to originate from 8-oxoG mispairing with A during replication. Furthermore, rapid cell divisions seemed to be required for fixation of mutations, as a similar dose of radiation did not increase the mutation frequency, or the frequency of GC to TA transversion, in the adult brain.

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